The overall objective of the proposed research is to understand the role of oxygen tension in determining coronary microvascular tone through the modulation of endothelium-dependent vasomotion. Low oxygen tension, or hypoxia, is an important component of a complex series of events occurring during ischemia and reperfusion that affects vascular function. The goal of the research is to relate changes in microvessel endothelial cell function to the events that occur pathologically during myocardial hypoperfusion. Specific aims of the research are to 1) determine the relationship between EDRF, nitric oxide, and vasodilator prostanoid activity during hypoxia. Previously, we demonstrated that oxygen tension alters both EDRF and prostanoid activity in microvessels, and hypoxia decreases EDRF activity. We now have evidence that inhibition of EDRF synthesis results in increased vasorelaxation by prostanoids released in response to hypoxia. Using an in vitro microvessel technique and tissue culture methods the research will determine if EDRF influences prostanoid activity by altering either the synthesis or release of prostaglandins by the endothelium. 2) determine the effects of hypoxia in vivo on endothelium-dependent vasodilation. Using an in vitro microvessel imaging technique to study vessels previously exposed to hypoxia while in the beating heart, the experiments will determine the effects of hypoxia on endothelium-dependent vasomotion in microvessels obtained from a specific coronary vascular bed perfused selectively in vivo with blood or Krebs saline pre-equilibrated to low oxygen tension with an extracorporeal oxygenator in the acute anesthetized dog model. The research will specifically focus on hypoxia-induced changes in EDRF and vasodilator prostanoid activity and will extend our previous studies on microvessels directly exposed to varying oxygen tensions in vitro. Microvessels will be studied in vitro to assess endothelium-dependent and independent responses, and compared to microvessels isolated from a control vascular bed. Microvascular responses will be determined in microvessels perfused with either whole blood or saline free of formed blood elements, complement, and circulating humoral agents. 3) ascertain if an endothelium-derived constricting factor is released by the coronary microcirculation in response to hypoxia. In large epicardial coronary vessels and peripheral vessels, low oxygen tension elicits the release of endothelium-derived constricting factors. Using pharmacologic methods, we plan to determine if the specific constricting factor endothelin is released in microvessels in response to hypoxia, and determine if there is a functional relationship between endothelium- derived vasoconstrictor activity and EDRF activity. The proposed research has direct application to the clinical problems of unstable angina and myocardial infarction. It will provide insight into alterations in the complex mechanisms responsible for the local control of microvascular tone and coronary blood flow that occur during ischemia/reperfusion.